Melanoma is one type of skin cancer referred to as malignant melanoma. There are various types of skin cancer. Melanoma is the type of skin cancer that has the highest grade of malignancy. Among cells composing skin, melanin-pigment-producing cells are referred to as pigment-producing cells (melanocytes). These cells become cancerous and melanoma is developed.
The frequency of the occurrence of melanoma in Japan is approximately 1.5 to 2 people per population of 100,000. It is inferred that melanoma annually occurs among approximately 1,500 to 2,000 people in Japan. In the Europe and the United States, the frequency of melanoma incidence is said to be over a dozen people per population of 100,000. The frequency of melanoma incidence in Australia is 20 or more people per population of 100,000, which is said to be the highest in the world. Accordingly, people in Europe, the United States and Australia are interested in melanoma and pay attention to its occurrence. Surprisingly, it has been confirmed that the frequency of the occurrence of melanoma is increasing yearly in both Japan and countries other than Japan. In the latest survey, the annual number of fatalities due to this disease in Japan was as high as around 450 people. Melanoma afflicts people regardless of age. Melanoma occurs more frequently particularly among people aged 40 and up and occurs most frequently occurs among people aged 60 to 70 and up. Melanoma occurs infrequently among children, but this does not mean that no melanoma occurs among children. Currently, melanoma tends to occur more frequently among young people aged 20 to 30 and up. There are no particular tendencies for melanoma to occur more frequently either in males or females. Melanoma occurs in both males and females. Melanomas in Japanese people are most frequently developed on the soles of the feet (planta pedis). Such melanoma cases account for approximately 30% of all melanoma cases in Japan. Melanomas in Japanese people are characterized in that they are often developed in the toenails and fingernails. Furthermore, similar to melanomas in Western people, melanomas in Japanese people are developed at all skin locations, such as in the trunk, hands, feet, face, head, and the like.
Serum tumor marker measurement is important not only in melanoma diagnosis, but also in early detection of recurrence in postoperative cases and in determination of therapeutic effects in cases at progressive stages. As tumor markers for melanoma, the usefulness of serum LDH and of 5-S-cysteinyldopa (5-S-CD) have been known to date. In more recent years, an S-100β protein and a melanoma inhibitory activity (MIA) have been reported as more sensitive markers. In Japan, 5-S-CD is broadly used as a tumor marker for melanoma. However, none of these tumor markers yield positive results unless the melanoma tested for is at a highly advanced stage, such as Stage IV. Therefore, it must be said that the usefulness of these tumor markers is limited in terms of melanoma diagnosis and early detection of postoperative recurrence.
The present inventors have previously examined the expression profiles of 23,040 types of genes in 20 types of primary hepatocellular carcinoma (HCC) and in various normal organs, including those in prenatal periods, with the use of a genome-wide cDNA microarray containing such genes. As a result, the present inventors have discovered that glypican-3 (GPC3) is highly expressed in most HCC types, whereas GPC3 is expressed in the liver, kidney, and lungs during the prenatal period and is hardly expressed in an adult's normal organs other than the placenta. Moreover, the present inventors have reported that the glypican-3 (GPC3) is a secretory protein, that GPC3 can be detected in the sera of 40% of HCC patients using the ELISA method, and that GPC3 is useful as a novel tumor marker for HCC (Nakatsura T. et al., Biochem. Biophys. Res. Commun. 306, 16-25 (2003)).
In 1996, Pilia et al. reported that glypican-3 (GPC3) encoding one member of glypican family is mutated in Simpson-Golabi-Behmel syndrome (SGBS) patients (Pilia G et al., Nat. Genet. 12, 241-247 (1996)). SGBS is an X-linked disorder characterized by pre- and postnatal overgrowth and wide-ranging clinical expression, ranging from a very mild phenotype in female carriers to lethal symptoms in male infants (Neri G et al., Am. J. Med. Genet. 79, 279-283 (1998)). Examples of clinical features of SGBS include distinct facial appearance, cleft palate, ankylodactylia, hyperdactylia, accessory nipples, cystic disease of kidney, renal dysplasia, and congenital heart disease (Behmel A et al., Hum. Genet. 67, 409-413 (1984); Garganta C. L. and Bodurtha J. N., Am. J. Med. Genet. 44, 129-135 (1992); Golabi M and Rosen, L., Am. J. Med. Genet. 17, 345-358 (1984); and Gurrieri F et al., Am. J. Med. Genet. 44, 136-137 (1992)). It has been reported that most GPC3 mutations are point mutations or deletions of several nucleotides including exons (Hughes-Benzie R. M. et al., Am. J. Med. Genet. 66, 227-234 (1996); Lindsay S. et al., J. Med. Genet. 34, 480-483 (1997); Veugelers. M. et al., Hum. Mol. Genet. 9, 1321-1328 (2000); and Xuan J. Y. et al., J. Med. Genet. 36, 57-58 (1999)). Moreover, because of the lack of correlation between a patient's phenotype and a mutation location, SGBS may be induced by the deletion of a functional GPC3 protein or by other genetic factors relating to the relevant phenotype within a family and between families (Hughes-Benzie R. M. et al., Am. J. Med. Genet. 66, 227-234 (1996)). A study on GPC3-deficient mice has also supported this hypothesis (Cano-Gauci D. F. et al., J. Cell Biol. 146, 255-264 (1999)). Such mice have some abnormalities that are observed in SGBS patients, such as overgrowth, cystic disease of kidney, and renal dysplasia.
It has been reported that based on studies using Northern blot analysis, GPC3 mRNA is overexpressed in HCC, placenta, fetal liver, fetal lungs, and fetal kidney (Zhu Z. W. et al., Gut 48, 558-564 (2001); Hsu. H. C. et al., Cancer Res. 57, 5179-5184 (1997); and Pellegrini M. et al., Dev Dyn. 213, 431-439 (1998)).
Moreover, it has been reported that since GPC3 is a cell proliferation inhibitor and can induce apoptosis in a type of tumor cells (Cano-Gauci D. F. et al., J. Cell Biol. 146, 255-264 (1999); and Duenas Gonzales A. et al., J. Cell Biol. 141, 1407-1414 (1998)), GPC3 expression is down-regulated in tumors derived from various origins. Lin et al. have demonstrated that GPC3 is expressed in the normal ovary, but cannot be detected in specific types of ovarian cancer cell lines (Lin H. et al., Cancer Res. 59, 807-810 (1999)). In all cases where no GPC3 expression was observed, the GPC3 promoter was hypermethylated, no mutations were found in the coding regions, and GPC3 expression was recovered through treatment with a demethylating agent. It has been further reported that the ectopic expression of GPC3 inhibits colony-forming activity in several types of ovarian cancer cell lines. Other data that relates GPC3 to cancer are data obtained by a differential mRNA display study concerning normal rat mesothelial cells and mesothelioma cell lines (Murthy S. S. et al., Oncogene 19, 410-416 (2000)). In this study, it was discovered that GPC3 is always downregulated in tumor cell lines. Furthermore, similar downregulation has also been observed in primary rat mesothelioma- or human mesothelioma-derived cell lines. Similar to the cases of ovarian cancer, no mutations have been discovered in the GPC3-encoding sequence; however, abnormal methylation in the GPC3 promoter region has been observed in most cell lines. As reported (Duenas Gonzales A. et al., J. Cell Biol. 141, 1407-1414 (1998)), it has been demonstrated that the ectopic expression of GPC3 in mesothelioma cell lines inhibits colony-forming activity. Furthermore, Xiang et al. have recently reported that GPC3 is also not expressed in human mammary cancer (Xiang Y. Y. et al., Oncogene 20, 7408-7412 (2001)). These data suggest that GPC3 can act as a negative regulator in growth of these types of cancer. Specifically, GPC3 expression is reduced during tumor advancement in a type of cancer arising from adult GPC3-positive tissue. Furthermore, such reduction may play some roles in the development of a malignant phenotype.
In contrast, in the case of HCC, a tumor arises from a liver tissue wherein GPC3 is expressed only in fetal stage. Moreover, GPC3 expression tends to appear again at the time of transformation to malignancy. It is unclear whether or not such re-expression of GPC3 is important for advancement of such tumor. For the last few years, it has been revealed that cell surface heparan sulfate proteoglycan (HSPG) is required for the optimal activity of a heparin-binding growth factor such as a fibroblast growth factor (FGF) or Wnt (Yayon A. et al., Cell 64, 841-848 (1991); and Schlessinger J. et al., Cell 83, 357-360 (1995)). Glypicans constitute a GPI-anchored cell surface HSPG family. It is thus inferred that tissue-specific differences in the relation between tumorigenesis and GPC3 expression level are due to the fact that GPC3 regulates growth and survival factors differently in each tissue. GPC3 probably functions as a carcinoembryonic protein at least in these organs. It is not generally thought that a carcinoembryonic protein plays an important role in tumor advancement. However, the carcinoembryonic protein has been used as a tumor marker or a target of immunotherapy (Coggin J. H. Jr., CRC Critical Reviews in Oncology/Hematology 5, 37-55 (1992); and Matsuura H. and Hakomori S.-I., Proc. Natl. Acad. Sci. USA 82, 6517-6521 (1985)). Whether or not the carcinoembryonic behavior of GPC3 can be used for clinical applications and whether or not the re-expression of such glypican is important for HCC advancement remain to be studied.